The invention relates generally to methods and systems for adjusting the contrast of individual dyes in color images of materials comprising a plurality or mixtures of dyes.
Color adjustment by manipulating the red, green, and blue components is a common tool used in digital image processing and analysis, including application in which biological images are digitized, processed and analyzed. For example, hematoxylin and eosin (H&E) staining is the most common staining method in histology, and has been used over a hundred years. The H&E has been favored due to its low cost, fast preparation, easy image acquisition, and the extensive knowledge and training behind it. Hematoxylin is a dye extracted by boiling the wood of logwood trees found mainly in South America and West India. The active dye in hematoxylin staining is hematein which is oxidized hematoxylin rather than the hematoxylin itself. Hematoxylin is usually mixed with metallic salts also called mordants, to enhance the blue color. Hematoxylin stains nucleic acids such as chromatin in the nuclei and ribosomes with blue-purple. The most common counterstain of hematoxylin is eosin. The most common type of eosin dye in the literature is eosin Y. Eosin stains the cytoplasm, collagen, muscle fibers, extracellular structures and red blood cells. An H&E stain may have some yellow, brown colors for the pigments in the tissue.
The color of the H&E varies across different labs, manufacturers, and preparation. Some pathologists prefer a darkly stained background where some prefer a clean background with clear and sharp nuclear staining. The most important factors that effect the H&E color during the preparation are the dye concentration, mordant ratio, pH, oxidation and time.
Given such variations in staining techniques and pathogists' preferences, it would appear to be advantageous to be able to digitally and selectively adjust the intensity of individual stains in a given image. However, separating individual stains in an image of a biological material that has been stained with a mixture of, or multiple, stains, and then manipulating the stain images has not previously been addressed in the literature or by competitors.